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A eukaryotic initiation factor 5C is upregulated during metamorphosis in the cotton bollworm, Helicoverpa armigera.

Dong DJ, Wang JX, Zhao XF - BMC Dev. Biol. (2009)

Bottom Line: Ha-eIF5C expression was upregulated by 20-hydroxyecdysone (20E).Furthermore, the transcription of Ha-eIF5C was down regulated after silencing of ecdysteroid receptor (EcR) or Ultraspiracle protein (USP) by RNAi.These results suggested that during metamorphosis of the cotton bollworm, Ha-eIF5C was upregulated by 20E through the EcR and USP transcription factors.

View Article: PubMed Central - HTML - PubMed

Affiliation: The Key Laboratory of Plant Cell Engineering and Germplasm Innovation of Ministry of Education, School of Life Sciences, Shandong University, Jinan 250100, Shandong, PR China. dujuandong@yahoo.com.cn

ABSTRACT

Background: The orthologs of eukaryotic initiation factor 5C (eIF5C) are essential to the initiation of protein translation, and their regulation during development is not well known.

Results: A cDNA encoding a polypeptide of 419 amino acids containing an N-terminal leucine zipper motif and a C-terminal eIF5C domain was cloned from metamorphic larvae of Helicoverpa armigera. It was subsequently named Ha-eIF5C. Quantitative real-time PCR (QRT-PCR) revealed a high expression of the mRNA of Ha-eIF5C in the head-thorax, integument, midgut, and fat body during metamorphosis. Immunohistochemistry suggested that Ha-eIF5C was distributed into both the cytoplasm and the nucleus in the midgut, fat body and integument. Ha-eIF5C expression was upregulated by 20-hydroxyecdysone (20E). Furthermore, the transcription of Ha-eIF5C was down regulated after silencing of ecdysteroid receptor (EcR) or Ultraspiracle protein (USP) by RNAi.

Conclusion: These results suggested that during metamorphosis of the cotton bollworm, Ha-eIF5C was upregulated by 20E through the EcR and USP transcription factors.

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Immunocytochemical localization of eIF5C in the midgut. Panels A-C are negative controls with pre-immune rabbit serum; panels D-F are midgut from feeding 5th instar larva (5th-24 h), molting 5th instar larva (5th-HCS) and 6th-96 h (wandering) larva; panels G and J, H and K, I and L are the magnified D, E, F, respectively; nuclear staining was done by DAPI (G, H, I) and the positive signals were detected by ALEXA 488 assay (J, K, L), panels A-F are overlay. LM, lumen of midgut; LPC, larval polyploidy cells; ISC, intestinal stem cell; IMC, imaginal cells; Co, clumnar cells; Go, goblet cells. Scale bar = 100 μm.
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Figure 8: Immunocytochemical localization of eIF5C in the midgut. Panels A-C are negative controls with pre-immune rabbit serum; panels D-F are midgut from feeding 5th instar larva (5th-24 h), molting 5th instar larva (5th-HCS) and 6th-96 h (wandering) larva; panels G and J, H and K, I and L are the magnified D, E, F, respectively; nuclear staining was done by DAPI (G, H, I) and the positive signals were detected by ALEXA 488 assay (J, K, L), panels A-F are overlay. LM, lumen of midgut; LPC, larval polyploidy cells; ISC, intestinal stem cell; IMC, imaginal cells; Co, clumnar cells; Go, goblet cells. Scale bar = 100 μm.

Mentions: To verify the expression and localization of Ha-eIF5C, we performed an immunohistochemical analysis of the midgut (Fig. 8), fat body and integument (Fig. 9) from feeding 5th larvae (5th-24 h), molting 5th larvae (5th-HCS) and wandering 6th-96 h larvae (6th-96 h). In the 5th-HCS stage, the midgut epithelium consisted of larval polyploid cells (LPC, including columnar and goblet cells) and intestinal stem cells (ISC) (Fig. 8-K). Larval ISCs are the progenitors of the adult midgut epithelium. The larval polyploid cells moving into the lumen from the basement membrane were replaced by proliferating and differentiating ISCs at 6th-96 h (Fig. 8-L). At this point, groups of imaginal cells began to form cell layers. Our immunohistochemical analysis shows that Ha-eIF5C was distributed into both the cytoplasm and nucleus in the midgut during the feeding 5th, molting 5th and wandering 6th-96 h stages. Relatively strong fluorescent signals were detected on the outer peripheries of the midgut epithelium from worms during 5th instar feeding and molting stage, as well as the larval polyploid cells and the imaginal cells from larvae at wandering stage. Likewise, ISCs, muscle cells and basement membrane were localized in this area.


A eukaryotic initiation factor 5C is upregulated during metamorphosis in the cotton bollworm, Helicoverpa armigera.

Dong DJ, Wang JX, Zhao XF - BMC Dev. Biol. (2009)

Immunocytochemical localization of eIF5C in the midgut. Panels A-C are negative controls with pre-immune rabbit serum; panels D-F are midgut from feeding 5th instar larva (5th-24 h), molting 5th instar larva (5th-HCS) and 6th-96 h (wandering) larva; panels G and J, H and K, I and L are the magnified D, E, F, respectively; nuclear staining was done by DAPI (G, H, I) and the positive signals were detected by ALEXA 488 assay (J, K, L), panels A-F are overlay. LM, lumen of midgut; LPC, larval polyploidy cells; ISC, intestinal stem cell; IMC, imaginal cells; Co, clumnar cells; Go, goblet cells. Scale bar = 100 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2667495&req=5

Figure 8: Immunocytochemical localization of eIF5C in the midgut. Panels A-C are negative controls with pre-immune rabbit serum; panels D-F are midgut from feeding 5th instar larva (5th-24 h), molting 5th instar larva (5th-HCS) and 6th-96 h (wandering) larva; panels G and J, H and K, I and L are the magnified D, E, F, respectively; nuclear staining was done by DAPI (G, H, I) and the positive signals were detected by ALEXA 488 assay (J, K, L), panels A-F are overlay. LM, lumen of midgut; LPC, larval polyploidy cells; ISC, intestinal stem cell; IMC, imaginal cells; Co, clumnar cells; Go, goblet cells. Scale bar = 100 μm.
Mentions: To verify the expression and localization of Ha-eIF5C, we performed an immunohistochemical analysis of the midgut (Fig. 8), fat body and integument (Fig. 9) from feeding 5th larvae (5th-24 h), molting 5th larvae (5th-HCS) and wandering 6th-96 h larvae (6th-96 h). In the 5th-HCS stage, the midgut epithelium consisted of larval polyploid cells (LPC, including columnar and goblet cells) and intestinal stem cells (ISC) (Fig. 8-K). Larval ISCs are the progenitors of the adult midgut epithelium. The larval polyploid cells moving into the lumen from the basement membrane were replaced by proliferating and differentiating ISCs at 6th-96 h (Fig. 8-L). At this point, groups of imaginal cells began to form cell layers. Our immunohistochemical analysis shows that Ha-eIF5C was distributed into both the cytoplasm and nucleus in the midgut during the feeding 5th, molting 5th and wandering 6th-96 h stages. Relatively strong fluorescent signals were detected on the outer peripheries of the midgut epithelium from worms during 5th instar feeding and molting stage, as well as the larval polyploid cells and the imaginal cells from larvae at wandering stage. Likewise, ISCs, muscle cells and basement membrane were localized in this area.

Bottom Line: Ha-eIF5C expression was upregulated by 20-hydroxyecdysone (20E).Furthermore, the transcription of Ha-eIF5C was down regulated after silencing of ecdysteroid receptor (EcR) or Ultraspiracle protein (USP) by RNAi.These results suggested that during metamorphosis of the cotton bollworm, Ha-eIF5C was upregulated by 20E through the EcR and USP transcription factors.

View Article: PubMed Central - HTML - PubMed

Affiliation: The Key Laboratory of Plant Cell Engineering and Germplasm Innovation of Ministry of Education, School of Life Sciences, Shandong University, Jinan 250100, Shandong, PR China. dujuandong@yahoo.com.cn

ABSTRACT

Background: The orthologs of eukaryotic initiation factor 5C (eIF5C) are essential to the initiation of protein translation, and their regulation during development is not well known.

Results: A cDNA encoding a polypeptide of 419 amino acids containing an N-terminal leucine zipper motif and a C-terminal eIF5C domain was cloned from metamorphic larvae of Helicoverpa armigera. It was subsequently named Ha-eIF5C. Quantitative real-time PCR (QRT-PCR) revealed a high expression of the mRNA of Ha-eIF5C in the head-thorax, integument, midgut, and fat body during metamorphosis. Immunohistochemistry suggested that Ha-eIF5C was distributed into both the cytoplasm and the nucleus in the midgut, fat body and integument. Ha-eIF5C expression was upregulated by 20-hydroxyecdysone (20E). Furthermore, the transcription of Ha-eIF5C was down regulated after silencing of ecdysteroid receptor (EcR) or Ultraspiracle protein (USP) by RNAi.

Conclusion: These results suggested that during metamorphosis of the cotton bollworm, Ha-eIF5C was upregulated by 20E through the EcR and USP transcription factors.

Show MeSH